Anorexia nervosa (AN) is a psychiatric illness with no accepted pharmacological treatment [1] and with one of the highest mortality rates among the mental illnesses (10-20%) [2-4]. Even if not fatal, AN can cause life-long damages to multiple organ systems, creating an increased burden on the health care system. AN has a stereotypical age of onset at puberty, with 90-95% of cases occurring in females [5]. This developmental pattern suggests that hormonal changes in females associated with puberty may trigger changes in brain connections that increase an individual's vulnerability to stress, anxiety and AN. AN is also associated with frequent relapses[6], suggesting that anorectic behavior during this pivotal, final stage of brain development may cause long-lasting changes in brain connections. The long-term goal of this project is to identify developmental changes at central synapses that are linked to AN vulnerability among females entering puberty and to characterize changes that are induced further by this illness and following recovery. We will test a novel hypothesis - namely, that pubertal females are more vulnerable to AN due to fluctuation in the release of a neurosteroid, THP, that triggers an increased expression of a4b2d GABAA receptors at the plasma membrane of hippocampal pyramidal neurons which, in turn, renders the hippocampus hyper-excitable during stressful events. This THP-GABA hypothesis will be tested by using an animal model of AN, activity-based anorexia (ABA), which captures multiple key features of AN but for which the sex- and age-dependence have not been fully explored. We will first run behavioral tests to determine whether the two key factors that influence the human population - age and sex - also influence ABA vulnerability of rodents. This will be achieved by comparing the behavioral factors related to the development of ABA, recovery from ABA and ABA relapse across 3 developmental stages (prepubertal, pubertal, and adulthood) and 2 sexes (male and female). We will then use biochemical and electron microscopic immunocytochemical approaches to determine whether a4b2d GABAA receptor expressions in certain brain regions (biochemical data) and at synapses (EM data) correlate with ABA vulnerability, development, recovery and relapse. To further test the THP-GABA hypothesis, we will also determine whether ABA vulnerability and EM/Biochemical changes associated with the onset of puberty among females are reduced by pre-treatments that target the THP-GABA system. PUBLIC HEALTH RELEVANCE: Anorexia nervosa (AN) is a psychiatric illness occurring predominately among females entering puberty, with one of the highest mortality rates among mental illnesses (10-20%) and no accepted pharmacological treatment. The aim of this proposal is to test a novel hypothesis - namely, that females entering puberty are more vulnerable to AN because the limbic system of female brains at this stage in development express more of a particular type of neurotransmitter receptor that is sensitive to both GABA (an inhibitory neurotransmitter) and a stress-related hormone, THP (also called allopregnanolone). The results obtained from this study will provide the rationale for exploring a new pharmacologic treatment that targets the site of action of THP upon the GABAergic system within limbic pathways.